1. Field of the Invention
This invention is directed to a class of compounds which are cocaine analogs, and which inhibit catecholamine, ago endoleamine, octopamine and phenylethanolamine reuptake transporters. Specifically, a novel family of compounds shows high binding specificity and activity, and can-be used to bind to these transporters. Such binding, and the inhibition of the transporter which results, is useful for controlling invertebrate pests, including inhibiting the feeding of and delaying the maturation of the invertebrate pests. The compounds also have a reduced ability to cross the blood-brain barrier in animals, and thus, the compounds are safe for use as insecticides.
2. Description of the Related Art
In mammals, the CNS stipulatory and euphoric effects of cocaine are thought to be due to cocaine's well-documented action in blocking dopamine (DA) reuptake into presynaptic nerve terminals (Jaffe, J., in The Pharmacological Basis of Therapeutics, Gilman, A. G., et al. , eds., MacMillan: New York, pp. 535-584 (1980); Kennedy, L., et al., J. Neurochem. 41: 172-178 (1983) Kuhar et al. 1991). Because amine reuptake is a major mechanism for inactivation of DA following its synaptic release, the effect of cocaine is to augment and prolong DA neurotransmission. In man, with moderate amounts of cocaine, this results several physiologic and psychologic effects (Jaffe 1980).
Pharmacological evidence from vertebrates supports the presence of distinct reuptake sites (amine transporter proteins) for DA, NE, and serotonin (5-HT) (Ritz, M., et al., NIDA Research Monograph 95: 239-246 (1989)). Although much recent emphasis has been put on cocaine's action on DA, older literature clearly indicates that cocaine also blocks the reuptake of NE and 5-HT (Baldessarini, R., et al., J. Neurochem. 18: 2519-2533 (1971); Body, T., et al. , Pharmacol. Biochem. & Behavior 34: 165-172 (1989)).
Early biochemical studies of monoamine reuptake in mammals involved the use of living animals injected with labeled amines or the use of intact, isolated organs or tissue fragments incubated in vitro with tracer (Hertting et al., J. Pharmacol. Exp. Ther. 134: 146-153 (1961); Dengler et al., Nature 191: 816-817 (1961); Axelrod and Inscoe, J. Pharmacol. Exp. Ther. 141: 161-165 (1963)). Although these studies yielded important data about reuptake kinetics, they have been less useful for pharmacologically characterizing transporter binding sites and for determining the structure-activity relationships of drugs capable of blocking these sites. This is due to the fact that drug penetration through intact tissues and drug metabolism within tissues can significantly alter measurement of the true affinity of compounds for transporter binding sites. Studies with intact tissue are also tedious and complicate the use of replicate samples because of the tube-to-tube variability in the size and homogeneity of the intact tissue pieces. Thus, many researchers have abandoned the use of intact tissue preparations in the characterization of vertebrate amine transporters and have used, instead, broken cell tissue fractions prepared in such a way as to contain intact pinched-off nerve endings (synaptosomes) capable of accumulating labeled amines under sodium- and energy-dependent conditions (Baldessarini and Vogt, J. Neurochem 18: 2519-2533 (1971); Anderson, J. Neurochem 48: 1887-1896 (1987); Boja and Kuhar, Europ. J. Pharmacol. 173: 215-217 (1989). Although cocaine has a fascinating medicinal history in man, dating back at least 4500 years, its natural function in plants is unknown (Plowman, T., in Ethnobotany in the Neotropics, France, G. T., et al., eds., N.Y. Botanical Garden, Bronx, N.Y., pp. 62-111 (1984)). Plowman, Rivier and others (Rivier, L., J. Ethnopharmacology 3: 313-335 (1981); Plowman, T., et al., Ann. Bot. (London) 51: 641-659 (1983)) have determined that the four major varieties of Erythroxylum (coca) plants that produce cocaine, contain levels ranging from 0.35-0.72% dry weight, with values often exceeding 1% (particularly in small, newly emerging leaves). Although relatively little is known about the insect pests of coca, Plowman & Well (J. Ethnopharmacology 1: 263-278 (1979)) have commented, on the basis of personal observations, that, compared with other tropical American crops, E. coca and E. novogranatense are relatively pest-free. Herbivorous insects are only rarely observed on the plants in the field; damage to leaves is often minor. This is especially noteworthy since, during much of the year, the membranaceous leaves of coca are found in the tender state of unfolding, the result of their being stripped 3-6 times a year during harvest.
Octopamine (OA) is an invertebrate-specific neurotransmitter which was first discovered over 45 years ago in the posterior salivary gland of the octopus (V. Erspamer and G. Boretti, Arch. Int. Pharmaco. Ther. 88: 926-322 (1951)). Although similar to norepinephrine (NE) in structure, OA has very little activity as a sympathomimetic when injected into mammals (A. Lands and J. Grant, J. Pharm. Exptl. Therap. 106: 341-345 (1952)) and, compared with NE, is present in very low concentrations in vertebrate tissues (Y. Kakimoto and M;. Armstrong, J. Biol. Chem. 237: 422-427 (1962)). Relatively little attention was paid to OA until early 1970's, when Molinoff and Axelrod reported that OA was present in much higher concentrations in invertebrates, particularly in invertebrate nerve tissue (P. B. Molinoff and J. Axelrod, J. Neurochem. 19: 157-163 (1972)).
In 1973, the first identification of an OA receptor was reported (J. A. Nathanson "Cyclic AMP: A Possible Role in Insect Nervous System Function", (Ph.D. Thesis) (1973); J. A. Nathanson and P. Greengard, Science, 19: 308-310 (1973)). Because this receptor was present in highest concentrations in insect nerve cord, it was postulated that OA might function as a neurotransmitter. Furthermore, because these receptors were undetectable in mammalian tissues, it was also postulated that the neurotransmitter function of OA might be largely restricted to invertebrates (J. A. Nathanson "Cyclic AMP: A Possible Role in Insect Nervous System Function", (Ph.D. Thesis) (1973); J. A. Nathanson and P. Greengard, Science, 19: 308-310 (1973); J. A. Nathanson, Trace Amines and the Brain: Eds. Marcel Dekker, pp. 161-190 (1976)). At about the same time, Kravitz and coworkers (B. Wallace et al., Brain Res. 349-55 (1974)) independently reported the presence of OA-containing neurons in crustacea, and, somewhat later, Hoyle reported evidence suggesting the presence of large OA neurons in insect ganglia (G. Hoyle, J. Exp. Zool 193: 425-31 (1975)). Subsequent work by a number of investigators has established the role of OA, not only as a neurotransmitter, but also as a neuromodulator and circulating neurohormone in insects and acarines (for review see I. Orchard, Can. J. Zool, 60: 659-69 (1982); H. A. Robertson and A. V. Juorio, Int. Rev. Neurobiol. 19: 173-224 (1976)). Indeed, OA plays a pervasive role in regulating many areas of insect physiology, including carbohydrate metabolism, lipid mobilization, hematocyte function, heart rate, peripheral muscle tension and excitability, and behavior. The functions that OA carries out in insects appear analogous to those carried out by norepinephrine (NE) and epinephrine (EPI) in vertebrates. This has led to the suggestion that, during evolution, there may have been a divergence in the use of these amines between the two arms of the animal kingdom (H. A. Robertson and A. V. Juorio, Int. Rev. Neurobiol. 19: 173-224 (1976); A. V. Robertson and A. V. Juorio, J. Neurochem. 28: 573-79 (1977); J. A. Nathanson, Physiological Reviews 57: 158-256 (1977)).
Analogous to the action of NE and EPI in vertebrates, many of the effects of OA in invertebrates are mediated by cyclic AMP (J. A. Nathanson and P. Greengard, Science 19: 308-310 (1973); J. A. Nathanson, Physiological Reviews 57: 158-256 (1977); H. Robertson and J. Steele, J. Neurochem 19: 1603-06 (1972); A. Harmar and A. Horn, Mol. Pharmacol. 13: 512-20 (1976); C. Lingle et al., Handbook of Exptl. Pharmacology, (J. Kebabian & J. Nathanson, eds.), pp. 787-846 (1982)). OA stimulates production of cyclic AMP through activation of OA-sensitive (G, protein-coupled) adenylate cyclase (J. A. Nathanson, J. Cyclic Nucleotide and Protein Phosphor. Res. 10: 157-66 (1985)). In 1979, it was found that the firefly light organ, in which OA mediates neural control of light emission (A. D. Carlson, Advances Insect Physiol. 6: 51-96 (1969); J. F. Case and L. G. Strause, Bioluminescence in Action (P. J. Herring, ed.), pp. 331-366 (1978)), has a virtually pure population of OA receptors present in enormous quantity, with no evidence of adenylate cyclases activated by other hormones (J. A. Nathanson, Science 203: 65-8 (1979); J. A. Nathanson and E. Hunnicutt, J. Exp. Zool. 208: 255-62 (1979a)). Thereafter, the first detailed pharmacological characterization of G.sub.s -linked OA receptors was carried out in the absence of other amine receptors (J. A. Nathanson, Science 203: 65-8 (1979); J. A. Nathanson and E. Hunnicutt, J. Exp. Zool. 208: 255-62 (1979a); J. A. Nathanson, Proc. Natl. Acad. Sci. U.S.A. 82: 599-603 (1985b); Nathanson, J. A., in Insect Neurochemistry and Neurophysiology, Borkovec, A., et al., eds., Humana: Clifton, N.J., pp. 263-266 (1986); Nathanson, J. A., et al., Neurosci. Abstr. 5:346 (1979)). More recently, a new chemical class of potent OA receptor agonists has been characterized, the phenyliminoimidazolidines (PIIs) (Nathanson, J. A., Proc. Natl. Acad. Sci. U.S.A. 82: 599-603 (1985); Nathanson, J. A., Mol. Pharmacol. 28: 254-268 (1985)). With the PIIs and other compounds, it has been possible to distinguish clearly the characteristics of OA receptors from those of mammalian adrenergic, dopaminergic, and serotonergic receptors.
Overactivation of the OA system in insects and acarines leads to behavioral and physiological abnormalities that have pestistatic and pesticidal consequences. One way to cause OA overactivation, and thereby take advantage of this system for pesticide development, is to directly stimulate OA receptor proteins.
Analogous to the octopamine neurotransmitter system is the cholinergic system, where the plant alkaloid nicotine exerts natural pesticidal effects through excessive activation of acetylcholine (ACh) receptors. As is well known, for pesticide development it has turned out that, more effective than cholinergic agonists, are the reversible and irreversible acetylcholinesterase inhibitors. Acetylcholinesterase (AChE) catalyzes the hydrolysis of the neurotransmitter ACh to choline and acetate. If AChE is inhibited by a pesticide, normal inactivation of ACh is blocked, and ACh accumulates to abnormally high levels. This causes overactivation of ACh receptors, indirectly, through inhibition of neurotransmitter degradation. It has recently been discovered that an analogous site of action exists for the OA system. Because of OA's selectivity for invertebrates, agents affecting this site will have reduced toxicity for vertebrates.
Because OA affects so many sites in insects, it is not surprising that disruption of this system adversely affects insect physiology. In 1980, it was reported that the formamidine pesticides cause glowing of firefly light organs and it was suggested that these compounds, which have low toxicity for vertebrates, might be exerting their pesticidal actions by affecting OA receptors. Subsequently, several labs (Nathanson, J. A., et al., Molec. Pharmacol. 20: 68-75 (1981); Evans, P. D., et al., Nature 287: 60-62 (1980)), determined that the formamidines are indeed potent OA agonists in several insect species. In addition, it was found that OA itself, as well as OA analogs and the PIIs applied to leaves, could markedly interfere with the feeding of M. sexta (Nathanson, J. A., Proc. Natl. Acad. Sci. U.S.A. 82: 599-603 (1985); Nathanson, J. A., in Insect Neurochemistry and Neurophysiology, Borkovec, A., et al., eds., Humana: Clifton. N.J., pp. 263-266 (1986); Nathanson, J. A., Mol. Pharmacol. 28: 254-268 (1985); Nathanson, J. A., in Sites of Action for Neurotoxic Pesticides, Hollingworth. R., et al., eds., Am. Chem. Soc.: Washington, D.C., pp. 154-161 (1987): Nathanson, J. A., Science 226: 184-187 (1984); Nathanson, J. A., in Abstr. 2nd Internatl. Symp. Insect Neurobiol. Pest. Action, Society of Chemical Industry: London, pp. 129-130 (1985); Nathanson, J. A., in Membrane Receptors and Enzyme as Targets of Insecticidal Action, Clark, J., et al., Plenum: New York, pp. 157-171 (1986)). The behavioral and pestistatic effects of these compounds on Manduca were similar to those of the formamidines: they caused tremors, hyperactivity, rearing, and poor coordination (resulting in leaf drop-off), abnormalities which, interestingly, are reminiscent of the effects of overdoses of amphetamines and adrenergic agonists in vertebrates.
Additional support for a connection between overactivation of the OA system and pesticidal activity has come from observations showing that the known species variation in the pesticidal effects of formamidines (Matsumura, F., et al., Environ. Health Perspect. 14: 71-82 (1976)) is related to the ability of these compounds to activate G.sub.s -coupled OA receptors (G.sub.s -coupled receptors are those whose activation results in the stimulation of adenylate cyclase and the synthesis of cyclic AMP). For example, in Manduca, a sensitive species, it has been found (Nathanson, J. A., Mol. Pharmacol. 28: 254-268 (1985)) that didemethylchlordimeform (DDCDM) is a full OA agonist, 20-fold more potent than OA, while in cockroach, a resistant species, DDCDM is much weaker than OA in activating adenylate cyclase. This species variability appears to result from the distribution of OA receptor subtypes that need to be specifically targeted for pesticide activity. (Additional evidence for involvement of G.sub.s (cAMP-linked) OA receptors comes from the observations (Nathanson, J. A., Proc. Natl. Acad. Sci. U.S.A. 82: 599-603 (1985) that the antifeeding effects of OA agonists are enhanced by inhibitors of cAMP catabolism and mimicked by adenylate cyclase activators (forskolin) and lipid-soluble cAMP analogs.)
In insects, uptake studies of monoamines have been largely limited to the study of octopamine, where evidence from intact tissue experiments supports the presence of a high affinity sodium-dependent transporter (Evans, J. Neurochem 30: 1015-1022 (1978); Carlson and Evans, J. Exp. Biol. 122: 369-385 (1986); Wierenga and Hollingworth, J. Neurochem. 54: 479-489 (1990)). These studies have utilized either intact ventral nerve cord or intact pieces of tissue incubated in insect saline. Surprisingly, there appears to be no report of the examination of the uptake of octopamine or other monoamines in synaptosomal-containing broken cell fractions. This appears to be the case despite the fact that several reports have described preparations of both crude and purified synaptosomal fractions from insects (Breer and Jererich, Insect Biochem. 10: 457-463 (1980); Whitton et al., Biochem. Soc. Trans. 14: 609-610 (1986); Luo and Bodnaryk, Insect Biochem. 17: 911-918 (1987); Nicholson and Connelly, Insect Biochem. 21: 447-456 (1991)).